ABSTRACT Infection with Zika virus (ZIKV), first linked with an increased incidence of microcephaly in Brazil, is now associated with a spectrum of fetal and postnatal malformations and developmental disorders. Our previous studies with rhesus macaques have demonstrated that, like the human population, severe fetal outcomes are uncommon, but they have also revealed that vertical transmission of ZIKV to the fetus may more common than is currently appreciated, with 100% transmission to fetuses when dams were inoculated in the first or third trimester. Additional neurological deficits from intrauterine infection are now becoming revealed in postnatal life, thus a better understanding of the correlates of vertical transmission at the maternal-fetal interface is critical. Investigators at the University of Wisconsin-Madison are funded to conduct a study to define the viral persistence and distribution in maternal tissues of pregnant rhesus macaques at staged intervals following infection with ZIKV. That study specifically will evaluate maternal tissues, but does not support concurrent evaluation of the maternal-fetal interface. We hypothesize that the antiviral response in the decidua and placenta can determine the risk for vertical transmission of the virus and the degree of fetal pathology and developmental dysregulation. To address this hypothesis, we propose to comprehensively define the immune response at the maternal-fetal interface in these pregnancies in two Specific Aims: Specific Aim 1. To define the T cell, antigen-presenting cell, and innate lymphoid/natural killer cell populations in control and ZIKV-infected decidua, establishing their transcriptional, secreted, and cell-surface profiles with high-dimensional flow cytometry and dimension reduction methodologies. Specific Aim 2. To define the Hofbauer cell surface, secreted, and activation profiles in control and ZIKV- infected placentas. We will use high-dimensional flow cytometric analytical methods established in our laboratories for human decidual leukocytes, guided by our previous work with rhesus decidual leukocytes and placental Hofbauer cells, comparing infected pregnancies with control pregnancies to be supported by this R21 proposal. The ZIKV- infected decidual and placental immunome will be interpreted in light of the distribution, burden, and pathogenic impact of ZIKV replication in the decidua and the fetus. A clearer understanding of the decidual and placental immune response with ZIKV infection, which we can couple with determination of fetal infection and pathology to be gained from the current study, will allow insight into the correlates of decidual and placental immune responses of fetal outcomes.